Ole Alvseike

Comparison between the growth of Yersinia enterocolitica, listeria monocytogenes, escherichia coli O157:H7 and salmonella spp. in ground beef packed by three commercially used packaging techniques.

Growth of the pathogens Yersinia enterocolitica, Listeria monocytogenes, Escherichia coli O157:H7 and strains of Salmonella were compared in ground beef packed in modified atmospheres of 60% CO2/40% N2/0.4% CO (high CO2/low CO mixture), 70% O2/30% CO2 (high O2 mixture) and in chub packs (stuffed in plastic casings). The ground beef was inoculated with rifampicin-resistant or nalidixic acid/streptomycin-resistant strains of the pathogens (final concentration 10(2) - 10(3) bacteria/g) and stored at 4 and 10 degrees C for up to 14 days. At 4 degrees C the shelf life, based on colour stability and background flora development, was prolonged for the high CO2/low CO mixture compared to the two other packaging methods, but at 10 degrees C the shelf life was < 8 days for all the packaging methods. Growth of Y. enterocolitica was nearly totally inhibited both at 4 and 10 degrees C in the high CO2/low CO mixture, while the bacterial numbers in the samples packed in the high O2 mixture increased from about 5 x 10(2) bacteria/g at day 0 to about 10(4) at day 5 at 4 degrees C and to 10(5) at 10 degrees C. Growth in the chub packs was even higher. L. monocytogenes showed very little growth at 4 degrees C in all treatments. At 10 degrees C there was slow growth from about 5 x 10(3) bacteria/g to about 10(4) at day 5 in the high CO2/low CO mixture, while the numbers in the high O2 mixture and the chub packs were about 10 times higher. Growth of E. coli O157:H7 at 10 degrees C in the ground beef was nearly totally inhibited in both the high CO2/low CO mixture and the high O2 mixture. Growth in the chub packs was higher, as the number of bacteria increased 3 log in 5 days. The Salmonella strains (S. typhimurium, S. dublin, S. enteritidis and S. enterica 61:k:1,5,(7)) in the ground beef stored at 10 degrees C for 5 and 7 days grew to a higher number in the high CO2/low CO mixture than in the high O2 mixture. This study shows that the growth of Y. enterocolitica and L. mononcytogenes in ground beef stored in the high CO2 /low CO mixture was not increased as a result of prolonging the shelf life. However, the observed growth of strains of Salmonella at 10 degrees C in this mixture and in chub packs does emphasise the importance of temperature control during storage.

Reduction of verotoxigenic Escherichia coli in production of fermented sausages

After a number of foodborne outbreaks of verotoxigenic Escherichia coli involving fermented sausages, some countries have imposed regulations on sausage production. For example, the US Food Safety and Inspection Service requires a 5 log(10) reduction of E. coli in fermented products. Such regulations have led to a number of studies on the inactivation of E. coli in fermented sausages by changing processing and post-processing conditions. Several factors influence the survival of E. coli such as pre-treatment of the meat, amount of NaCl, nitrite and lactic acid, water activity, pH, choice of starter cultures and addition of antimicrobial compounds. Also process variables like fermentation temperature and storage time play important roles. Though a large variety of different production processes of sausages exist, generally the reduction of E. coli caused by production is in the range 1-2 log(10). In many cases this may not be enough to ensure microbial food safety. By optimising ingredients and process parameters it is possible to increase E. coli reduction to some extent, but in some cases still other post process treatments may be required. Such treatments may be storage at ambient temperatures, specific heat treatments, high pressure processing or irradiation. HACCP analyses have identified the quality of the raw materials, low temperature in the batter when preparing the sausages and a rapid pH drop during fermentation as critical control points in sausage production. This review summarises the literature on the reduction verotoxigenic E. coli in production of fermented sausages.

Heterogeneity in resistance to food-related stresses and biofilm formation ability among verocytotoxigenic Escherichia coli strains.

This study assessed the resistance of ten verocytotoxigenic Escherichia coli (VTEC) isolates of commonly encountered serogroups/-types and two non-pathogenic E. coli strains to various food-related stresses (acid, alkaline, heat and high hydrostatic pressure treatments) and their biofilm formation ability. In addition, the global changes in the cellular composition in response to the exposure to these adverse environments were monitored by Fourier Transform Infrared (FT-IR) spectroscopy for two of the strains. Large inter-strain variations in stress resistance were observed. The most tolerant strains belonged to serogroup O157 which included both the O157:H7 type strain EDL933 and a representative isolate of the sorbitol fermenting O157:H- VTEC clone (strain MF3582). Strain C-600, a non-pathogenic laboratory strain, was sensitive to multiple stresses. Although wide variation in biofilm-forming ability was observed among VTEC isolates, no consistent relationships between biofilm-forming ability and capacity to withstand stress exposures were found. Analysis of the allelic status of the rpoS gene, involved in the general stress response of stationary-phase cells, allowed detection of loss-of-function mutations for two strains, E218/02 and MF2411, both of them showing as common features a high sensitivity to alkaline and heat treatments and a poor ability to form mature biofilms. Evidences found in this study confirm rpoS as a highly mutable gene in nature, and suggest its relevance not only for the mount of an active stress response but also for the establishment of mature biofilm communities. Our findings contribute to increase the knowledge on the resistance of VTEC to environmental stresses commonly encountered in the food chain, which can lead to improved strategies for preventing VTEC infections.

Application of high pressure processing to reduce verotoxigenic E. coli in two types of dry-fermented sausage.

The effect of high pressure processing (HPP) on the survival of verotoxigenic Escherichia coli (VTEC) in two types of Norwegian type dry-fermented sausages was studied. Two different types of recipes for each sausage type were produced. The sausage batter was inoculated with 6.8 log(10) CFU/g of VTEC O103:H25. After fermentation, drying and maturation, slices of finished sausages were vacuum packed and subjected to two treatment regimes of HPP. One group was treated at 600 MPa for 10 min and another at three cycles of 600 MPa for 200 s per cycle. A generalized linear model split by recipe type showed that these two HPP treatments on standard recipe sausages reduced E. coli by 2.9 log(10) CFU/g and 3.3 log(10) CFU/g, respectively. In the recipe with higher levels of dextrose, sodium chloride and sodium nitrite E. coli reduction was 2.7 log(10) CFU/g in both treatments. The data show that HPP has a potential to make the sausages safer and also that the effect depends somewhat on recipe.

Prevalence of a Salmonella subspecies diarizonae in Norwegian sheep herds

We studied the prevalence of Salmonella enterica subspecies diarizonae serovar 61:k:1,5,(7) (S. IIIb 61:k:1,5,(7)) in 133 Norwegian sheep herds from throughout the country. We used a nested case-control study to identify major risk factors. Two pooled samples (one from lambs and one from adult sheep) were collected from each herd from (10 different) abattoirs. The regional prevalence varied from 0 to 45%, and was more prevalent in adults than in lambs. Sixteen of the 133 herds tested positive to S. IIIb 61:k:1,5,(7), and these positive herds were included in the questionnaire-based nested case-control part of the study. The controls were stratified according to abattoir. Above-median herd size, having white crossbreed sheep, and higher annual replacement percentage were the most evident risk factors.

The prevalence and dynamics of Salmonella enterica IIIb 61:k:1,5,(7) in sheep flocks in Norway.

Fifty randomly selected sheep flocks from a region in central Norway were sampled in December 1999 to determine the flock prevalence of Salmonella enterica subspecies diarizonae serovar 61:k:1,5,(7) (S. IIIb 61:k:1,5,(7)). From each flock, 15-41 rectal swabs were collected from individual sheep of different age groups and examined for S. IIIb 61:k:1,5,(7). Positive flocks were visited again in January-April and each time, rectal swabs from the same animals were collected and examined for this specific serovar. Seven flocks (14%; 95% CI 6.3-27%) were positive for S. IIIb 61:k:1,5,(7) in December; in all, 10 sheep out of the 1233 (0.8%) were positive at the first sampling. From the seven positive flocks, six, five, six, and nine animals were positive in January, February, March, and April, respectively. Of the total 21 individual sheep tested positive from January to April, 15 were >2 years old (OR(ex)=3.26; 1.1-10.2). Six out of the seven positive flocks were large flocks (>117 ewes). Sharing of rams between flocks did not seem to be a risk factor for the presence of S. IIIb 61:k:1,5,(7) in a flock.

Effects of post-processing treatments on sensory quality and Shiga toxigenic Escherichia coli reductions in dry-fermented sausages

The effects of post-processing treatments on sensory quality and reduction of Shiga toxigenic Escherichia coli (STEC) in three formulations of two types of dry-fermented sausage (DFS; salami and morr) were evaluated. Tested interventions provided only marginal changes in sensory preference and characteristics. Total STEC reductions in heat treated DFS (32°C, 6days or 43°C, 24h) were from 3.5 to >5.5 log from production start. Storing of sausages (20°C, 1month) gave >1 log additional STEC reduction. Freezing and thawing of sausages in combination with storage (4°C, 1month) gave an additional 0.7 to 3.0 log reduction in STEC. Overall >5.5 log STEC reductions were obtained after storage and freezing/thawing of DFS with increased levels of glucose and salt. This study suggests that combined formulation optimisation and post-process strategies should be applicable for implementation in DFS production to obtain DFS with enhanced microbial safety and high sensory acceptance and quality.

Microbiological, physicochemical and sensory parameters of dry fermented sausages manufactured with high hydrostatic pressure processed raw meat

The aim of this trial was to describe physicochemical, microbiological and organoleptic characteristics of dry fermented sausages produced from high hydrostatic pressure (HHP) pre-processed trimmings. During ripening of the meat products pH, weight, water activity (aw), and several microbiological parameters were measured at zero, eight, fifteen days and after 6weeks. Sensory characteristics were estimated at day 15 and after six weeks by a test panel by using several sensory tests. Enterobacteriaceae were not detected in sausages from HHP-processed trimmings. Fermentation was little affected, but weight and aw of the HHP-processed sausages decreased faster during ripening. HHP-treated sausages were consistently less favoured than non HHP-treated sausages, but the strategy may be an alternative approach if the process is optimized.

Effects of hygienic treatments during slaughtering on microbial dynamics and contamination of sheep meat.

The aims of this study were to investigate bacterial dynamics in the sheep meat chain, from fleece to meat trimmings, using both quantitative and qualitative analyses, and to study the effects on microbial load associated with the hygienic interventions of: i) shearing sheep immediately before slaughter, ii) manual steam vacuum pasteurisation, iii) hot water pasteurisation of carcasses, followed by iv) chilling. A further aim was to provide evidence to determine whether or not unshorn sheep should be handled in a processing line separate from that of shorn sheep in Norwegian abattoirs. A total of 176 surface swab samples were collected from three sites along the value chain: i) on fleeces, ii) on carcasses at the end of the slaughter line, and iii) on carcasses after chilling for 24h, and 32 samples were collected from meat trimmings. The results showed that Aerobic Plate Counts (APC) were lower for the shorn group compared to the unshorn group, both on carcasses before chilling and after chilling (difference of 0.8 and 0.9logCFU/1000cm(2) (p≤0.05), respectively) and in meat trimmings (difference of 0.5logCFU/g (p≤0.05)). Hygienic treatments were used on carcasses derived from unshorn sheep, and steam vacuum treatment reduced Escherichia coli, Enterobacteriaceae, and APC before chilling by 1.2, 1.0, and 0.6logCFU/1000cm(2) (p≤0.05), respectively, and hot water pasteurisation, in addition to chilling, reduced E. coli, Enterobacteriaceae, and APC by 0.7, 1.0, and 0.9logCFU/1000cm(2) (p≤0.05), respectively, compared with untreated carcasses. The effect of chilling was shown by the significant reduction of number of carcasses where E. coli were detected; from 65% (13/20) of the shorn group before chilling to 35% (7/20) after chilling, and from 90% (36/40) to 45% (9/20) of the unshorn group. Sequencing of the 16S rRNA gene derived from 316 colonies of Enterobacteriaceae showed a tendency for the relative proportion of the genus Escherichia/Shigella, compared with other genera within Enterobacteriaceae, to be greater for unshorn, untreated sheep than from the other groups at the sampling locations along the meat chain. The study showed that steam vacuum and hot water pasteurisation reduced the contamination of carcasses derived from unshorn sheep, down to the level of the shorn group, and thus can replace the separate processing line for unshorn sheep. Indeed, the low microbial contamination in meat trimmings for all groups indicates that the separate processing line is unnecessary.

Growth of Salmonella choleraesuis subspecies diarizonae serovar 61:k:1,5,(7) in broth and fresh mutton.

Three serovars of Salmonella choleraesuis (IIIb 61:k:1,5,(7), Enteritidis and Dublin) were grown in broths of pH 5.5 and 6.2 and incubated at 4, 6, 8 or 12 degrees C. Growth in the broth, measured by means of an increase of absorbance, was not observed below 8 degrees C. At 8 and 12 degrees C, the maximum growth rate (mu(max)), lag period and maximum absorbance level (max(abs)) varied according to serovar and pH. In general, serovar IIIb 61:k:1,5,(7) and serovar Enteritidis grew better than serovar Dublin. The effect of pH on lag period, seen for serovar IIIb 61:k:1,5,(7) and serovar Enteritidis at 8 degrees C, was absent at 12 degrees C, while the effect of pH regarding the mu(max) and the max(abs) was observed also at 12 degrees C. Furthermore, the growth serovar IIIb 61:k:1,5,(7) in normal and dark, firm and dry meat at 8 degrees C with ambient air in competition with the natural microbial flora was tested in minced meat and chops. Slow growth of serovar IIIb 61:k:1,5,(7) was observed in minced meat. The low virulence and the ordinary growth capabilities indicate that serovar IIIb 61:k:1,5,(7) will probably not represent a serious hazard to the public health.